The enormous HIV-1 sequence diversity is a major obstacle to vaccine development. Numerous candidate vaccines representative of multiple strains are in the pipeline. Objective evaluation of candidate vaccines in clinical trials is essential for vaccine development and optimization. Our goal is to develop standardized peptide panels that can be used to objectively compare CD8+ T cell responses among candidate vaccines. Detection of greater magnitude and breadth of T cell responses in an HIV-1-infected population with a given candidate peptide panel will be surrogate for a superior candidate peptide panel to assess T cell responses to candidate vaccines in that population. Our studies will focus on two highly immunogenic regions of HIV-1, Gag p17 and Nef, which contain a spectrum of amino acid variability within them. The specific aims are to: 1. Assess the magnitude and breadth of clade-specific and cross-clade T cell responses in 40 subjects each with CRF02_AG (Senegal), subtype B (US), and subtype C (India) infections, using two central sequence [Consensus (Con) and Center of Tree (COT)] based peptide sets. We will perform rigorous testing of subtype A, B, and C peptides based on the Con and COT sequences by IFN-gamma ELISpot in three populations with different circulating forms that predominate the current global epidemic. 2. Compare T cell responses to Gag p17 and Nef peptide sets designed for optimal coverage of potential T cell epitopes (PTE-optimized) with responses to central sequence (Con and COT) peptide sets in 40 subtype B-infected subjects. We will test candidate peptide panels specifically designed to augment PTE coverage by IFN-gamma ELISpot. 2. Compare T cell responses to Gag p17 PTE-optimized peptides with responses to autologous peptides in 10 subtype B infected patients. We will determine autologous HIV-1 sequences encoding Gag p17 from the plasma of ten subtype-B infected subjects and examine T cell response to autologous peptides by IFN-gamma ELISpot. The findings will lead to the design of optimal peptide reagents for the objective evaluation and comparison of T cell responses to candidate HIV-1 vaccines in clinical trials. The studies will also provide insight into the design of vaccines with expanded T cell epitope coverage that will elicit broad T cell responses recognizing multiple epitope variants.